# Seed cultures in 6-well plates at ~2.5x10<suP>5</sup> so that cells will be ~5.0x10<sup>5</sup> at the time of transfection. Use 1.0 mL antibiotic-free growth medium (e.g., 10% FBS in McCoy’s 5A for, no pen/strep) per well.

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# Seed cultures in 6-well plates at ~2.5x10<suP>5</sup> so that cells will be ~5.0x10<sup>5</sup> at the time of transfection.

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* '''NOTE:''' if you are using a cell line that does not adhere strongly to the plate (eg, HEK cells), seed the cultures using antibiotic-free medium the day before to avoid having to replace medium the day of transfection.

Mammalian Cell Transfection - Lipofectamine LTX Reagent

Materials

Procedure - 6-well Format

1 day before transfection:

Seed cultures in 6-well plates at ~2.5x105 so that cells will be ~5.0x105 at the time of transfection.

NOTE: if you are using a cell line that does not adhere strongly to the plate (eg, HEK cells), seed the cultures using antibiotic-free medium the day before to avoid having to replace medium the day of transfection.

Transfection:

At your bench, bring 2.0 μg total plasmid DNA up to 20 μL total volume with fresh, sterile DNase-free water in fresh, sterile 1.5 mL tubes. Take the DNA samples to the tissue culture room.